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Objective To optimize the supercritical CO2 extraction conditions of volatile oil from Wenjing Huoxue cataplasm. Methods On the basis of single factor investigation on the comprehensive score of extraction yield , osthole content and isoimperatorin, the effects of extraction temperature, pressure and time on the comprehensive score of extracted volatile oil were optimized by orthogonal design. Results In the single factor experiment, the factors that had a great influence on the comprehensive score of the extracted volatile oil were extraction temperature, extraction pressure and extraction time. The orthogonal experiment results showed that the extraction temperature and extraction pressure had a significant influence on the comprehensive score of volatile oil. The optimized extraction process was as follows: extraction temperature at 55 ℃, extraction pressure as 30 MPa, and extraction time as 2 h. Conclusion The extraction process optimized in this experiment is stable and feasible, which could be used for the extraction and preparation of the volatile oil.
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Objective To improve the quality control of Dilong Shenmai oral liquid. Methods TLC was used for the qualitative identification of Astragali Radix, Ophiopogonis Radix and Schisandrae Chinensis Fructus in Dilong Shenmai oral liquid. HPLC was used to determine the contents of schisandrin and ethylparaben in the preparation. Wondasil C18 column (250 mm×4.6 mm, 5 μm) was used with acetonitrile-water as the mobile phase at the flow rate of 1.0 ml/min for gradient elution. The detection wavelength was set at 254 nm, and column temperature was 30 ℃. Results TLC spots were clear and well-separated without negative interference. The linear ranges of schisandrin and ethylparaben were 5.81−58.06 μg/ml (r=0.999 9) and 25.29−252.94 μg/ml (r=0.999 9). The average recoveries were 99.35% (RSD=1.02%) and 99.72% (RSD=0.76%). Conclusion This method is simple, quick and accurate. It can be used for effective quality control of Dilong Shenmai oral liquid.
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Objective To establish a HPLC fingerprints of Biyuanjing capsules. Methods The column was Agilent SB-C18(4.6mm×250 mm, 5 µm). The mobile phase was acetonitrile-water with gradient elution at a flow rate of 1.0 ml/min. The detection wavelength was 210 nm. The detection time was 80 min. Results The HPLC fingerprints of Biyuanjing capsules were established. Twenty common peaks were confirmed, of which, 15 peaks were belonging to each crude drug and 5 peaks were identified as chemical components. The overall similarity of the fingerprints of 10 batches of samples was above 90% comparing with the control. Conclusion This method can be used for the quality control of Biyuanjing capsules.
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OBJECTIVE： To study the anti-tumor effect of anemoside B4 （AB4） on hepatocellular carcinoma Huh-7 and tumor-bearing nude mice and its mechanism. METHODS： Huh-7 cells were divided into AB4 treatment group， negative control group （constant volume of culture medium） and positive control group （5-fluorouracil 50 μmol/L）. The inhibitory rate of Huh-7 cell proliferation was tested by MTT after treated with 0， 3， 6， 12， 25， 50， 100， 200， 400， 800， 1 600 μmol/L AB4 for 12， 24， 36， 48 h； IC50 were calculated. The number of clone cells was evaluated by clone formation tests after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h. The apoptosis rate of Huh-7 cells was tested by Annexin Ⅴ-FITC/PI double staining after treated with 50 μmol/L AB4 for 24 h. The expression of apoptotic proteins such as Bcl-2， Bax， Caspase-3， Cleaved Caspase-3 and Cleaved PARP were tested by Western blot after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h. Tumor-bearing nude mice were randomly divided into negative control group （normal saline）， positive control group （5-fluorouracil 50 mg/kg）， AB4 lwo-dose， medium-dose and high-dose groups （25， 50， 100 mg/kg）， with 3 mice in each group. They were given relevant medicine intraperitoneally， once a day， for consecutive 19 d. The growth of tumor was observed， and anti-tumor rate was also calculated. HE staining was used to observe the morphology of tumor cells. RESULTS： The inhibition rate of AB4 on Huh-7 cell proliferation increased with the increase of concentration of AB4， but it did not increase significantly after reaching 50 μmol/L； it increased with the increase of time， but it did not increase significantly after 24 h. The IC50 of AB4 was （56.76±1.756） μmol/L. Compared with negative control group， the number of clone cells was decreased significantly after Huh-7 cells were treated with 50 μmol/L AB4 for 24 h， while the protein expression of Bcl-2 was decreased significantly （P＜0.05）； the apoptotic rate， the protein expression of Bax， Caspase-3， Cleaved Caspase-3 and Cleaved PARP were increased significantly （P＜0.05 or P＜0.01）， there was no statistical significance， compared with positive control group. Compared with negative control group， the volume of tumor was decreased significantly in AB4 low-dose， medium-dose and high-dose groups， positive control group （P＜0.05）； the outline of tumor cells become more and more blurred； there were nuclear pyknosis， unclear nucleoplasm and nuclear fragmentation； its anti-tumor rates were 25.93％， 39.15％， 46.26％， 42.83％， respectively. CONCLUSIONS： AB4 inhibits Huh-7 cells and tumor-bearing mice， the mechanism of which may be associated with up-regulating the proportion of Bax/Bcl-2， activating Caspase-3， cracking PARP and inducing apoptosis.
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Objective: To establish the HPLC fingerprints of compound Yinchen granules. Methods: The column was Agilent SB-C18(250 mm×4. 6 mm, 5 μm) and the mobile phase was acetonitrile (A)-0. 2% phosphoric acid solution (B) with gradient elution at a flow rate of 1. 0 ml·min-1. The column temperature was 25℃. The detection wavelength switching technology was used in 180-mi-nute elution time. Results: The HPLC fingerprints of compound Yinchen granules were established. Twenty-two common peaks were confirmed, of which five peaks were identified and 18 peaks were assigned to each crude drug. The overall similarity of the fingerprints of 10 batches of samples was 0. 9 or more when compared with the control map. Conclusion: The fingerprints of compound Yinchen granules can provide reference for the overall quality control of compound Yinchen granules.
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OBJECTIVE:To establish a method for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.METHODS:The dual-wavelength HPLC method was adopted.The determination was performed on Wondasil C18 column with mobile phase consisted of acetonitrile-0.1％ phosphoric acid (11∶89,V/V) at the flow rate of 1.0 mL/min.The detection wavelengths were 245 nm for (R,S)-goitrin and 327 nm for chlomgenic acid.The column temperature was 30 ℃,and injection volume was 10 μL.RESULTS:The linear ranges were 4.05-40.51 μg/mL for (R,S)-goitrin (r=0.999 9),29.41-294.05 μg/mL for chlorogenic acid (r=0.999 9).The limits of quantification were 3.32,2.45 ng,limits of detection were 1.00,0.74 ng.RSDs of precision,stability and reproducibility tests were lower than 1.0％;the recoveries were 98.46％-101.06％ (RSD=0.98％,n=9) and 98.18％-100.78％ (RSD=0.86％,n=9).CONCLUSIONS:The method is simple,accurate,sensitive and reproducible,and can be used for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.
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OBJECTIVE:To establish a method for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.METHODS:The dual-wavelength HPLC method was adopted.The determination was performed on Wondasil C18 column with mobile phase consisted of acetonitrile-0.1％ phosphoric acid (11∶89,V/V) at the flow rate of 1.0 mL/min.The detection wavelengths were 245 nm for (R,S)-goitrin and 327 nm for chlomgenic acid.The column temperature was 30 ℃,and injection volume was 10 μL.RESULTS:The linear ranges were 4.05-40.51 μg/mL for (R,S)-goitrin (r=0.999 9),29.41-294.05 μg/mL for chlorogenic acid (r=0.999 9).The limits of quantification were 3.32,2.45 ng,limits of detection were 1.00,0.74 ng.RSDs of precision,stability and reproducibility tests were lower than 1.0％;the recoveries were 98.46％-101.06％ (RSD=0.98％,n=9) and 98.18％-100.78％ (RSD=0.86％,n=9).CONCLUSIONS:The method is simple,accurate,sensitive and reproducible,and can be used for simultaneous determination of (R,S)-goitrin and chlorogenic acid in Banchai oral liquid.
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Objective:To investigate the preventive effect of tetramethylpyrazine (TMP) on hypoxic pulmonary hypertension in rats and the underlying mechanism.Methods:Thirty-six male Sprague-Dawley rats were randomly divided into three groups:the normal control group,hypobaric and hypoxic group and TMP group (100 mg·kg-1·d-1).After the animal models of hypobaric and hypoxic pulmonary hypertension were established,mean pulmonary artery pressure (mPAP),mean carotid artery pressure (mCAP),ratio of right ventricular/(lift ventricular + interventricular septum) (RVHI) and morphological changes of pulmonary vessels were observed and the rates of wall thickness/external diameter (WT%) and wall area/total vascular area (WA%) were calculated.The contents of nitric oxide (NO),endothelin-1 (ET-1),hypoxic-inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) were detected in serum after the 21-day treatment with TMP.Results:The values of mPAP,RVHI,WT% and WA% of hypobaric and hypoxic group were significant higher than those of the normal control group (P0.05).The values of NO in serum of hypobaric and hypoxic group was lower than those of the normal control group (P<0.05) and those of TMP group were obviously higher than those of hypobaric and hypoxic group (P<0.05).The values of ET-1,HIF-1α and VEGF in serum of hypobaric and hypoxic group were higher than those of the normal control group (P<0.05) and those of TMP group were obviously lower than those of hypobaric and hypoxic group (P<0.05).Conclusion:TMP can effectively prevent pulmonary hypertension induced by hypobaric and hypoxic and structural remodeling of pulmonary arterioles.The mechanism may be related to the content up-regulation of NO and activity down-regulation of ET-1,HIF-1α and VEGF in serum of rats.
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Objective:To establish the methods for the qualitative identification and content determination of aurantio-obtusin and chrysophanol in Zeju Jiangzhi tablets. Methods:A TLC method was adopted for the qualitative identification, and an HPLC method was used for the content determination. The determination was performed on a Wondasil C18 (250 mm × 4. 6 mm, 5 μm ) column with the mobile phase of acetonitrile -0. 1% phosphonic acid with gradient elution at the flow rate of 1. 0 ml?min-1 , the detection wave-length was 286 nm, the column temperature was 30℃ and the injection volume was 10μl. Results:The TLC spots of aurantio-obtusin and chrysophanol were clear and well-separated without any negative interference. The HPLC experiment results showed the good line-arity within the range of 1. 03-25. 72μg?ml-1(r=0. 999 9) for aurantio-obtusin, and 0. 48-11. 92μg?ml-1(r=0. 999 9) for chry-sophanol. The average recovery was 99. 21% and 98. 85%, and RSD was 0. 70% and 0. 73%, respectively (n=9). Conclusion:The method is simple, accurate and repeatable, which can be used for the qualitative identification and content determination of auran-tio-obtusin and chrysophanol in Zeju Jiangzhi tablets.
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OBJECTIVE To investigate the effect of the dosing time on the pharmacokinetics of gefitinib and its potential mechanism. METHODS Female BALB/c nude mice were housed under standardized 12 h light/dark circadian conditions(light on at 7:00,off at 19:00)for two weeks before a non-small cell lung cancer(NSCLC)model was established. Two weeks later,they were divided into 2 groups(8:00,20:00)randomly. Gefitinib was orally administered at the dose of 1 mg · kg-1 to the mice in each group at 8:00 or 20:00,respectively. Blood was collected at 10 different time points after each administration. Livers were collected every 4 h during the 24 h period from the non-administrated nude mice of NSCLC model. The plasma concentration of gefitinib was determined through an HPLC-MS/MS and the parameters were calculated by WinNonlin 6.3. The total RNA was extracted from livers,purified, synthesized to cDNA that was subjected to qRT-PCR analysis for mRNA expression levels of cyto?chrome P450 enzymes(Cyp)3a11,Cyp3a13,pregnane X receptor(PXR)and constitutive androstane receptor (CAR). RESULTS The area under the plasma concentration-time curve (AUC) and mean residence time (MRT) of 8:00 administration group were higher than those of 20:00 administration group(P<0.05). The clearance(Clz/F) of 8:00 administration group was lower than that of 20:00 administration group(P<0.05). The mRNA expression levels of PXR and CAR were consistent with those of Cyp3a11 and Cyp3a13. CONCLUSION Circadian rhythm exists in the pharmacokinetics of gefitinib and it may be closely related to CYP3A and its regulator genes.
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Objective:To investigate the protective effects of Panax notoginseng on myocardial ischemia injury in dogs and study the spectrum-activity relationship of Panax notoginseng. Methods:Firstly, the HPLC fingerprint analytical method for Panax notogin-seng was established, and then the dog model of acute myocardial ischemia was established by left anterior descending coronary liga-tion. Bivariate correlation analysis and multivariate regression analysis were used to correlate the spectrum-activity relationship between the fingerprints and the anti-myocardial ischaemia activity, and the spectrum-activity relationship and efficacy material foundation of Panax notoginseng were determined. Results:The main effective components were Ginseng saponin Rg1 and Rb1 and notoginseng sapo-nins R1 etc. Notoginseng saponins R1 could significantly inhibit the increase of serum lactate, and ginseng saponin Rg1 could inhibit the increase of FFA in serum, which was the main component in Panax notoginseng for the treatment of myocardial ischemia. Conclusion:The effective substances in Panax notoginseng are obtained by investigating the relationship between the spectrum and efficiency, and a new method for the evaluation of spectrum-activity relationship for Panax notoginseng is established. It can objectively reflect the inher-ent quality of the drug and provide a new strategy for the further research of traditional Chinese medicines.
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Biomarkers of non-small cell lung cancer(NSCLC)can facilitate an efficient diagnosis, improve therapeutic effects,reduce the side effects of other treatments,ameliorate prognosis and predict relapse of the disease and drug resistance. This paper summarized the biomarkers which were newly discovered and their functions in clinical practices in order to guide individualized treatment and help adjust treatment. The exploration of biomarkers of NSCLC will become a priority in future research.
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To review and analyze the pharmacological effect and underlying mechanism of Chinese lobelia. Methods:The references from home and abroad on the pharmacological effect and mechanism of Chinese lobelia in recent years were summarized and analyzed. Results:Chinese lobelia had wide range of pharmacological effects, such as anti-tumor, endothelial cells regulation, an-algesic and anti-inflammatory effect, inhibition against alpha glycosidase enzymes and myocardial ischemia reperfusion et al. Conclu-sion:A large number of effective compositions are in Chinese lobelia and the biological activity is extensive. The mechanism of biologi-cal activity should be further studied in-depth.
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Objective:To re-establish the quality control standard for Weiling granules. Methods:The 4 chief herbs in the prepa-ration, radices paeoniae alba, licorice, rhizoma corydalis and hawthorn were identified by TLC qualitatively. The content of paeoniflor-in in radices paeoniae alba was determined by HPLC. The separation was performed on an Agilent XDB-C18 (250 mm × 4. 6 mm, 5μm)coulme with mobile phase consisting of acetonitrile-0. 1% phosphoric acid solution (10:90). The detection wavelength was 230 nm and the flow rate was 1. 0 ml·min-1 . Results:The spots in TLC were clear without any interference. The linear range for paeoni-florin was 0. 151-1. 212 μg(r=0. 999 9,n=5). The average recovery was 99. 63% and RSD was 2. 01%(n=9). Conclusion:The method is simple and accurate with high reproducibility, which can be used for the quality control of Weiling granules.
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OBJECTIVE:To establish the quality standard for Yiwei granule. METHODS:TLC was conducted for the qualitative identification of Coptis chinensis,Corydalis yanhusuo and Schisandra chinensis;HPLC was conduced for the content determination of paeoniflorin. The column was XDB C18 with mobile phase of methanol-water(25:75,V/V)at flow rate of 1.0 ml/min,detection wave-length was 230 nm and injection volume was 20μl. RESULTS:The TLC showed clear spots and good separation. The linear range of paeoniflorin was 7.575-60.6 μg/ml(r=0.999 9);RSDs of precision,stability and reproducibility tests were lower than 2.0%,recov-ery was 96.97%-102.51%(RSD=1.92%,n=9). CONCLUSIONS:The method is simple and reproducible,and can be used for the quality control of Yiwei granule.
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OBJECTIVE:To optimize the water-extraction technology of Compound kuhuang formulation. METHODS:The wa-ter-extraction technology of Compound kuhuang formulation was optimized by orthogonal design,with the total contents of matrine and oxymatrine,the total content of prim-O-glucosylcimifugin and 5-O-methylvisamminol,the content of berberine hydrochloride and extract yield,and with the extraction time,amount of water(3 reflux extractions)and soaking time as the factors;and verifi-cation tests were conducted. RESULTS:The extraction time had significant effect on comprehensive evaluation value (P<0.05). The optimal water-extraction technology was as follows as extraction time of 120 min,water amount of 8,6,and 6 times as the amount of herbs,and soaking time of 20 min. In verification tests,the average total content of matrine and oxymatrine was 3.152 6 mg/g(RSD=1.03%,n=3),that of prim-O-glucosylcimifugin and 5-O-methylvisamminol was 4.977 2 mg/g(RSD=2.27%,n=3),the average content of berberine hydrochloride was 3.345 0 mg/g (RSD=1.19%,n=3) and the average extract yield was 49.23%(RSD=2.43%,n=3). CONCLUSIONS:The optimal water-extraction technology of Compound kuhuang formulation is stable and feasible.
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Objective To establish the standard for quality control ofQingyan Granule. Methods The chief components of the preparation, Sophora Tonkinensis radix et rhizoma, Adenophorae radix, Lonicera japonica caulis, and Ophiopogonis radix were identified by TLC qualitatively. The contents of licorice glycosides and glycyrrhizic acid were determined by HPLC. The separation was performed on Thermo Syncronis C18 column (4.6 mm×250 mm, 5μm) with mobile phase consisted of acetonitrile with 0.05% phosphoric acid solution (A)-0.05% phosphoric acid solution (B), and gradient elution (0-8 min, 19%A;8-35 min, 19%→50%A). Detection wavelength was 237 nm, and flow rate was 1 mL/min.Results The spots in TLC were clear. There were spots with same color on the corresponding location of reference substance and reference herbal, negative control without interference. The linear range for licorice glycosides was 0.05-0.5μg (r=0.999 9). The average recovery was 99.97%, RSD=1.74% (n=9). The linear range for glycyrrhizic acid was 0.1-2μg (r=0.999 9). The average recovery was 99.74%, RSD=1.28% (n=9). Conclusion The method is simple, accurate, with high reproducibility, which can be used for quality control ofQingyan Granule.
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<p><b>OBJECTIVE</b>To explore the expression of MMP2 and its correlation with the clinical features and prognosis of endometrial adenocarcinoma.</p><p><b>METHODS</b>We collected paraffin-embedded samples from 81 patients with endometrial adenocarcinoma aged 32 to 80 years to examine the expression of MMP2 using immunohistochemistry. The correlation of MMP2 expression with the clinical characteristics and prognosis of the patients was analyzed.</p><p><b>RESULTS</b>MMP2 protein was expressed in the cytoplasm of the tumor cells. MMP2 over-expression was negatively correlated with tumor differentiation (P=0.015) and prognosis (P=0.041) of endometrial adenocarcinoma.</p><p><b>CONCLUSION</b>MMP2 over-expression is a potential malignant biomarker in endometrial adenocarcinoma.</p>
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Adult , Aged , Aged, 80 and over , Female , Humans , Middle Aged , Biomarkers, Tumor , Metabolism , Endometrial Neoplasms , Metabolism , Pathology , Matrix Metalloproteinase 2 , MetabolismABSTRACT
OBJECTIVE To investigate the effect of the dosing time on the pharmacokinetics oferlotinib.METHODS Female C57BL mice were contained under standardized 12h light/dark circadianconditions (lights on at 7:00,off at 19:00)for 3 weeks and randomly assigned into six groups.Erlotinibhydrochloride was orally administrated to the mice in each group at 8:00,12:00,16:00,20:00,24:00and 4:00,respectively.Blood was drawn from the eyeballs of the mice at 12 different time points aftereach administration.The plasma concentration of erlotinib was determined through a high-performanceliquid-chromatographic assay and the parameters were calculated by WinNonlin. RESULTSThe area under curre (AUC)and mean residence time (MRT)of these groups were apparently differ-ent.AUC0 ~24 h and MRT0 ~24 h were the lowest in the 20:00 administration group as compared to othergroups (P<0.01 ).Tmax of the 20:00,24:00 and 4:00 groups was apparently higher than that of the8:00 and 12:00 groups (P<0.01 ).The clearance of the light phase groups was lower than that of thedark phase groups (P<0.01 ),with the highest in the 20:00 group.The peak value of cmax appeared inthe 12:00 group and the lowest in the 20:00 group (P<0.01 ).CONCLUSION Circadian rhythm playsa critical role in pharmacokinetics of erlotinib in mice.
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Objective:To compare the steam distillation( SD) and supercritical CO2 ( SFE-CO2 ) method in the extraction of vola-tile constituents from such 3 traditional Chinese drugs as cnidium monnieri, radix sileris and atractylodes lancea in compound Kuhuang prescription. Methods:Essential oil was extracted from the 3 traditional Chinese drugs by SD and SFE-CO2 method, respectively. The components were identified by GC-MS, and their relative contents were calculated with peak area normalization method. Results:To-tally 36 components were isolated and identified using SD extraction and 31 components were identified using SFE-CO2 extraction, a-mong them, 22 components were the same and their relative molecular weight mainly concentrated within 200-230. Conclusion: SFE-CO2 method can extract effective components from the 3 traditional Chinese drugs in compound Kuhuang prescription with higher selec-tivity, which is the more suitable extraction method for essential oil from the prescription.